1,4,7,10-tetraazacyclododecane butyltriols, processes for their production and pharmaceutical agents containing them

ABSTRACT

1,4,7,10-Tetraazacyclodedecane butyltriols of general formula I A  
                 
 
in which
         R 1  means hydrogen or a metal ion equivalent independent of one another and   R 2  means a butyltriol radical   as well as their salts with organic or inorganic bases or amino acids are valuable pharmaceutical agents.

This is a continuation of U.S. application Ser. No. 09/359,045 filedJul. 23, 1999, which is a continuation of U.S. application Ser. No.07/671,041 filed Mar. 19, 1991 now U.S. Pat. No. 5,980,864.

BACKGROUND OF THE INVENTION

This invention relates to 1,4,7,10-tetraazacyclodedecane butyltriols,their complexes and complex salts, agents containing these compounds,their use as diagnostic agents and therapeutic agents as well asprocesses for the production of these compounds and agents.

In European patent application 87730085.5 with the publication number 0255 471 macrocyclic compounds of general formula I

are claimed,in which

Y is a nitrogen atom or phosphorus atom,

A¹ and A² are the same or different and each is a straight-chain orbranched alkylene group with 2 to 6 carbon atoms,

U¹, U², U³, U⁴ are the same or different and each is a direct bond or astraight-chain or branched alkylene group with 1 to 6 carbon atoms,

D¹, D², D³, D⁴ are the same or different and each is an oxygen atom orsulfur atom, an alkylene group with 1 to 6 carbon atoms or a group N-R⁷with R⁷ meaning a hydrogen atom, a straight-chain or branched alkylenechain with 1 to 4 carbon atoms, which on the end carries a COOR¹ group,and R¹ stands for a hydrogen atom or a metal ion equivalent.

D⁵ has the meaning indicated for D¹, D², D³ and D⁴ aswell as the group

with R⁵ meaning a hydrogen atom or a straight-chain or branchedsaturated or unsaturated C₁-C₂₀ alkylene group, optionally substitutedby hydroxy, mercapto, imino and/or amino group(s), optionally containingimino, phenylenoxy, phenylenimino, amide, ester group(s), oxygenatom(s), sulfur atom(s) and/or nitrogen atom(s), which on the endexhibits either a functional group or a macromolecule B bound by it,

s and t represent whole numbers from 0 to 5,

R² represents hydrogen, a straight or branched, saturated or unsaturatedalkyl, acyl or acylalkyl group with 1 to 16 carbon atoms optionallysubstituted by one or more hydroxy or lower alkoxy groups,

—CH₂—X—V with X meaning carbonyl, a straight-chain or branched-chainalkylene group with 0 to 10 carbon atoms, which optionally issubstituted by one or more hydroxy or lower alkoxy groups or astraight-chain or branched-chain alkylene group interrupted by oxygenatoms with 2 to 23 hydrocarbon atoms,

V meaning

of —COOR⁶, and R³ and R⁴, independently of one another, representhydrogen, a straight or branched alkyl group with 1 to 16 carbon atomsoptionally substituted by one or more hydroxy or lower alkoxy groups orR³ and R⁴ together with the nitrogen atom represent a saturated five orsix ring optionally containing another heteroatom and R⁶ representshydrogen or a saturated, unsaturated straight-chain or branched-chain orcyclic hydrocarbon radical with up to 16 carbon atoms or an aryl oraralkyl group,

or

R² or R³ represent a second macrocycle of formula I′—bound by analkylene chain (K) containing 2 to 20 carbon atoms, which optionallycarries carbonyl groups on the ends and optionally is interrupted by oneor more oxygen atoms or R¹ carboxymethylimino groups or is substitutedby one or more hydroxy, lower alkoxy or carboxy lower alkyl groups—

which can be of a different structure than the parent substance of thefirst, or

R² means B or CH₂—COB,

provided that, if R² stands for B or CH₂—COB, R⁵ means a hydrogen atom,that at least two COOR¹ groups are present in the molecule and that twoheteroatoms of the macrocycle are each connected by an alkylene groupwith at least two carbon atoms, and functional groups existing in themolecule optionally are conjugated with macromolecules and optionallyfree carboxyl groups are made into salts with organic or inorganic basesor amino acids and basic groups with inorganic or organic acids.

The substances and the solutions prepared from them meet the demands tobe made of pharmaceutically usable chelates. They have a strong andadaptable effectiveness, by the selection of suitable metal atoms, onthe respective principles of the diagnostic or therapeutic method (xray, NMR, ultrasound, nuclear medicine).

SUMMARY OF THE INVENTION

Of the many compounds in EP-A-0 255 471, the1,4,7,10-tetraazacyclodedecane butyltriols of general formula I_(A)

in which

R¹ means hydrogen or a metal ion equivalent independent of one anotherand

R² means a butyltriol radical

as well as their salts with organic or inorganic bases or amino acids,

exhibit such outstanding properties that even in comparison with thestructurally very closely related compound disclosed in EP-A 0 255 471(Example 6), the use of these selected compounds guarantees asubstantial advantage.

Compounds of general formula I_(A) with R¹ meaning hydrogen aredesignated as complexing agents and with at least two of substituents R¹meaning a metal ion equivalent are designated as metal complexes.

The element, which forms the central ion of the physiologicallycompatible complex salt, can, of course, also be radioactive for desiredpurpose of use of the diagnostic medium according to the invention.

If the medium according to the invention is intended for use in NMRdiagnosis, the central ion of the complex salt has to be paramagnetic.This involves especially the bivalent and trivalent ions of the elementsof the atomics numbers 21-29, 42, 44 and 58-70. Suitable ions are, forexample, the chromium(III), manganese(II), iron(II), cobalt(II),nickel(II), copper(II), praseodymium(III), neodymium(III), samarium(III)and ytterbium(III) ion. Because of their very strong magnetic momentthere are especially preferred the gadolinium(III), terbium(III),dysprosium(III), holmium(III), erbium(III) or iron(III) ion.

For use of the media according to the invention in nuclear medicine thecentral ion has to be radioactive. For example, radioisotopes of theelements copper, cobalt, gallium, germanium, yttrium, strontium,technetium, indium, ytterbium, gadolinium, samarium and iridium aresuitable.

If the medium according to the invention is intended for use in x-raydiagnosis, the central ion has to be derived from an element of higheratomic number to achieve a sufficient absorption of the x rays. It wasfound that for this purpose diagnostic media, which contain aphysiologically compatible complex salt with central ions of elements ofthe atomic numbers between 21-29, 42, 44, 57-83, are suitable; they are,for example, the lanthanum(III) ion and the above-named ions of thelanthanide series.

Preferred radicals R² are 2,3,4-trihydroxybutyl- and the1-hydroxymethyl-2,3-dihydroxypropyl radical.

If not all acidic hydrogen atoms are substituted by the central ion,one, several or all remaining hydrogen atom(s) can be replaced bycations of inorganic and/or organic bases or amino acids. Suitableinorganic cations are, for example, the lithium ion, the potassium ion,the calcium ion, the magnesium ion and especially the sodium ion.Suitable cations of organic bases are, among others, those of primary,secondary or tertiary amines, such as, for example, ethanolamine,diethanolamine, morpholine, glucamine, N,N-dimethylglucamine andespecially N-methylglucamine. Suitable cations of amino acids are, forexample, those of lysine, arginine and ornithine.

As proof for the above-named surprising and outstanding properties ofthe compounds according to the invention there are to be indicated theresults of animal experimental studies to determine the acuteintravenous (LD₅₀) as well as neural compatibility (ED₅₀) of twobutyltriol compounds according to the invention, i.e., of compounds ofgeneral formula I_(A) in comparison with the compound described in EP-A0 255 471, which structurally is closest to the two butyltriolmacrocycles:

1. Acute Toxicity Determination (LD₅₀)

In an individual cage (Rhema/Hofheim company) the contrast medium wasadministered to mice (weight: 18-22 g) approximately at body temperaturein a caudal vein at a rate of 2 ml/min. Also preferred are those of10-[2-hydroxy-2,2-bis(hydroxymethyl)ethyl]-1,4,7-triscarboxymethyl-1,4,7,10-tetraazacyclododecane,and10-[1,1,1-tris(hydroxymethyl)methyl]-1,4,7-triscarboxymethyl-1,4,7,10-tetraazacyclododecane.

The contrast media were administered to 3 mice each in 3 doses atvariable volumes and constant concentration. The allocation of theanimals to the individual doses and the injection sequence of the dosestook place by chance.

The observation period of the animals extended over 7 days perinjection. The death of the animals was used as the criterion of theeffectiveness.

2. Intracisternal Administration (ED₅₀)

The test substance was administered under light ether anesthesia to anequal number of female and male rats (130-170 g, Wistar-Han-Schering,SPF) once intracisternally by suboccipital puncture in constant volumes(0.08 ml/animal) in different concentrations (3 doses per 10 animals persubstance). A volume sampling check (Ringer's solution) followed thestudy. Posture anomalies, spasms and death of the animals were evaluatedas action criteria.

The statistical evaluation of all study results took place with theprobit analysis.

Neural Acute ED₅₀ Substance i.v. LD₅₀ (micromol/ (Gd complex) R²(mmol/kg) kg) Example 1 —CH₂CH(OH)—CH(OH)CH₂OH 35 27 of this inventionExample 2 —CH(CH₂OH)—CH(OH)—CH₂OH 30 29 of this invention Example 6—CH₂—CH(OH)—CH₂OH 25 14 EP-A 0255471

Contrast media for medical diagnosis should be eminently compatible andbehave in as biologically inert manner as possible. Monomer contrastmedia should come as close as possible to the intravenous compatibilityof, e.g., mannitol (i.e., 30-40 mmol/kg). Comparable compounds withlower values therefore are biologically not inert and show undesirableinteraction with the organism. The propanediol derivative from EP-A 0255 471 shows with an acute i.v. LD₅₀ of 25 mmol/kg clearly lower valuesthan the two compounds according to the invention. This relevantdifference indicates a certain chemotoxicity of the comparisonsubstance. The undesirable interaction was shown particularly in thecomparison of the test substances in the neural compatibility. The twocompounds according to the invention showed a clearly bettercompatibility. With the propanediol compound already at doses of 14micromol/kg changes in the behavior and marked spasms occurred. Someanimals died at this dose.

With the macrocyclic gadolinium complexes of this invention asubstantially better neural compatibility could be observed. Adetrimental effect on behavior occurred only with doses approximatelytwice as high as with the comparison substance.

Conclusion

Since the two butyltriol compounds according to the invention also showa lower osmolality (0.59 or 0.57 in comparison with 0.62 osmol/kg;aqueous solutions of the Gd complexes in a concentration of 0.5 mol/lwere measured) than the above-described propanediol compound, in summaryit can be noted that the undesirable interactions of this compound withthe biological organism surprisingly do not occur with the structurallyclosely related compounds according to the invention. The two butyltriolcompounds show marked advantages and therefore are substantially moresuitable as biologically inert contrast media.

The production of the compounds of general formula I_(A) according tothe invention takes place in that compounds of general formula II

in which

X stands for a nitrogen protecting group or CH₂COOY group with Y meaninghydrogen, an ammonium cation, an alkali metal or a protecting group,

are reacted with a substrate introducing the radical R² in protectedform, the optionally contained nitrogen protecting groups X are removedand the —NH— groups thus released are alkylated with an acetic acidderivative of general formula IIIHalCH₂COOY  (III)in which Hal stands for chlorine, bromine or iodine,

the (hydroxy and optionally acid) protecting groups are removed and theresulting compounds of general formula I_(A) with R¹ meaning hydrogenare converted with metal oxide or metal salt into the metal complexes ofgeneral formula I_(A) with R¹ meaning a metal ion equivalent and then—ifdesired—still existing acidic hydrogen atoms are substituted by cationsof inorganic and/or organic bases, amino acids or amino acid amides.

Suitable as acid protecting groups Y are lower alkyl, aryl and aralkylgroups, for example, the methyl, ethyl, propyl, n-butyl, t-butyl,phenyl, benzyl, diphenylmethyl, triphenylmethyl,bis(p-nitrophenyl)-methyl group as well as trialkylsilyl groups.

The acids HalCH₂COOH can also be used in the form of their salts,preferably as Na or K salt.

The cleavage of the acid protecting groups takes place according toprocesses known to one skilled in the art, for example, by hydrolysis,hydrogenolysis, alkaline saponification of the esters with alkali inaqueous alcoholic solution at temperatures of 0 to 50° C., acidsaponification with mineral acids or in case, e.g., of tert-butyl esterswith the help of trifluoroacetic acid.

The three nitrogen atoms of feedstock II carry, before the reactionintroducing the radical R², the group CH₂COOY or nitrogen protectinggroups, for example, the tosylate or trifluoroacetate group, which arecleaved according to methods known in the literature before thealkylation reaction that is to follow [the tosylates, e.g., with mineralacids, alkali metals in liquid ammonia, hydrobromic acid and phenol,RedAl^((R)), lithiumaluminum hydride, sodium amalgam, cf. e.g, LiebigsAnn. Chem. (1977), 1344, Tetrahedron Letters (1976), 3477; thetrifluoracetates, e.g. with mineral acids or ammonia in methanol,cf.e.g., Tetrahedron Letters (1967), 289].

The N-alkylation with a haloacetic acid derivative of general formulaIII takes place in polar aprotic solvents such as, for example,dimethylformamide, dimethyl sulfoxide, acetonitrile, aqueoustetrahydrofuran or hexamethylphosphoric acid triamide in the presence ofan acid trap such as, for example, tertiary amine (for example,triethylamine, trimethylamine, N,N-dimethylaminopyridine,1,5-diazabicyclo[4.3.0]-nonene-5(DBN),1,5-diazobicyclo[5.4.0]-undecene-5-(DBU), alkali, alkaline-earthcarbonate, bicarbonate or hydroxide (for example, lithium, sodium,magnesium, calcium, barium, potassium carbonate, hydroxide andbicarbonate) at temperatures between −10° C. and 120° C., preferablybetween 0° C. and 50° C.

Suitable as hydroxy protecting groups are all those that can easily beintroduced and later again be easily cleaved with re-formation of thefinally desired free hydroxy group. Preferred protecting groups areether groups such as, for example, the benzyl, 4-methoxybenzyl,4-nitrobenzyl, trityl, di- and tri-phenylmethyl, trimethylsilyl,dimethyl-t-butylsilyl, diphenyl-t-butylsilyl group. But preferably thehydroxy groups are protected in the form of ketals, for example, withacetone, acetaldehyde, cyclohexanone or benzaldehyde.

The cleavage of the hydroxy protecting groups takes place in a way knownin the art, for example, in case of a benzyl ether by reductive cleavagewith lithium/ammonia or by hydrogenolytic cleavage in the presence, forexample, of palladium-carbon and in case of a ether or ketal cleavage byacid treatment with the help, for example, of cation exchangers,trifluoroacetic acid or mineral acids [see, e.g., T. W. Greene“Protective Groups in Organic synthesis,” John Wiley and Sons (1981)].

The introduction of the radical R² takes place by alkylation of asubstrate synthesized from a nucleofuge group for example Cl, Br, I,CH₃C₆H₄SO₃, CF₃SO₃, CH₃SO₃ and of the protected radical R² or of asubstrate from which the desired radical R² is intramolecularlygenerated during the reaction. As an example for the last mentioned casethere can be mentioned, the hydroxy epoxides2,3-epoxy-1,4-dihydroxybutane and 1,2-epoxy-3,4-dihydroxybutaneprotected as, e.g., acetonides.

The reaction of feedstock II with X meaning a CH₂COOY group isperformed, e.g., in water, DMF, dioxane, alcohols, acetonitrile,tetrahydrofuran or their mixtures at temperatures of 0 to 100° C.,preferably room temperature to 60° C., at a basic pH, preferably 9 to13, within 6 hours to 2 days, preferably 12 to 36 hours.

If a macrocycle, protected on the residual nitrogen atoms, is used forthe introduction of the R² radical in the reaction, the reaction takesplace preferably in an autoclave in solvents such as, for example, DMF,DMA, toluene, methylene chloride or their mixtures at temperatures of 20to 170° C., preferably 100 to 150° C., with addition of a base, such as,e.g., amines, alkali, alkaline-earth hydroxides and carbonates,preferably potassium and sodium carbonate and hydroxide, within 6 hoursto 2 days, preferably 12 to 36 hours. If a substrate is used, whichcontains no nucleofuge group (i.e., for example, the above-mentionedepoxides), the use of a base can be dispensed with.

The compounds of general formula I_(A) with R¹ meaning a hydrogen atomrepresent complexing agents. They can be isolated and purified orwithout isolation can be converted into metal complexes of generalformula I_(a) with at least two of substituents R¹ meaning a metal ionequivalent.

The production of the metal complexes according to the invention takesplace in the way that was disclosed in German laid-open specification 3401 052, by the metal oxide or a metal salt (for example, the nitrate,acetate, carbonate, chloride or sulfate of the element of the atomicnumbers 21-29, 42, 44, 57-83) being dissolved or suspended in waterand/or a lower alcohol (such as methanol, ethanol or isopropanol) andbeing reacted with the solution or suspension of the equivalent amountof the complexing ligand and then, if desired, by existing acidichydrogen atoms being substituted by cations of inorganic and/or organicbases or amino acids.

The introduction of the desired metal ions take place both before andafter cleavage of the hydroxy protecting groups.

The neutralization of possibly still existing free carboxy groups takesplace with the help of inorganic bases (for example, hydroxides,carbonates or bicarbonates), for example, of sodium, potassium, lithium,magnesium or calcium and/or organic bases as, among others, primary,secondary and tertiary amines, such as, for example, ethanolamine,morpholine, glucamine, N-methyl and N,N-dimethylglucamine, as well asbasic amino acids, such as, for example, lysine, arginine and ornithineor of amides of originally neutral or acidic amino acids.

For the production of neutral complex compounds so much of the desiredbases is added to the acid complex salts in aqueous solution orsuspension that the neutral point is reached. The resulting solution canthen be evaporated to dryness in a vacuum. Often it is advantageous toprecipitate the formed neutral salts by addition of water-misciblesolvents, such as, for example, lower alcohols (methanol, ethanol,isopropanol and others), lower ketones (acetone and others), polarethers (tetrahydrofuran, dioxane, 1,2-dimethoxyethane and others) andthus obtain crystallizates that are easy to isolate and purify. It hasproved especially advantageous to add the desired base already duringthe complex formation of the reaction mixture and thus to save a processstep.

Another possibility to achieve neutral complex compounds consists inconverting the remaining acid groups in the complex entirely orpartially, for example, to esters or amides. This can occur bysubsequent reaction on the finished complex (e.g., by exhaustivereaction of the free carboxy groups with dimethyl sulfate.

The production of the pharmaceutical agents according to the inventionalso takes place in a way known in the art, by complex compoundsaccording to the invention—optionally by addition of additives usual ingalenicals—being suspended or dissolved in aqueous medium and then thesuspension or solution optionally being sterilized. Suitable additivesare, for example, physiologically safe buffers (such as, for example,tromethamine), additives of complexing agents (such as, for example,diethylenetriaminepentaacetic acid) or—if desired—electrolytes, such as,for example, sodium, calcium, magnesium, zinc chlorides, phosphates andcitrates or—if necessary—antioxidizing agents, such as, for example,ascorbic acid.

If suspensions or solutions of agents according to the invention inwater or physiological salt solution are desired for enteraladministration or other purposes, they are mixed with one or moreauxiliary agents usual in galenicals (for example, methylcellulose,lactose, mannitol) and/or surfactants (for example, lecithins,Tween^((R)), Myrj^((R))) and/or aromatic substance(s) for tastecorrection (for example, essential oils).

In principle it is also possible to produce pharmaceutical agentsaccording to the invention even without isolation of complex salts. Inany case, special care must be used to perform the chelate formation sothat the salts and salt solutions according to the invention areproduced practically free of uncomplexed toxically acting metal ions.

This can be guaranteed, for example with the help of color indicatorssuch as xylenol orange by control filtrations during the productionprocess. The invention therefore relates also to processes for theproduction of complex compounds and their salts. A purification of theisolated complex salt remains as final safety measure.

The pharmaceutical agents according to the invention preferably contain0.1 micromol-3 mol/l of the complex salt and generally are dosed inamounts of 0.1 micromol-5 mmol/kg. They are intended for enteral andparenteral administration. The complex compounds according to theinvention can be used:

1. for NMR and x-ray diagnosis in the form of their complexes with theions of elements without atomic numbers 21-29, 42, 44 and 57-83;

2. for radiodiagnosis and radiotherapy in the form of their complexeswith the radioisotopes of the elements with atomic numbers 27, 29, 31,32, 37-39, 43, 49, 62, 64, 70, 75 and 77.

The agents according to the invention meet the varied requirements forthe suitability as contrast media for nuclear spin tomography. Thus theyare outstandingly suitable for improving, in its expressive power, theimage obtained with the nuclear spin tomograph, by enhancing the signalintensity after enteral or parenteral administration. Further, they showthe great effectiveness, which is necessary, to load the body with thesmallest possible amounts of foreign substances, and the goodcompatibility, which is necessary to maintain the noninvasive characterof the examinations.

The good water solubility and low osmolality of the agents according tothe invention make it possible to produce highly concentrated solutions,to maintain the volume load of the circulation in justifiable limits andto balance the dilution by the body fluid, i.e., NMR diagnostic mediahave to be 100 to 1000 times better water soluble than for NMRspectroscopy. Further, the agents according to the invention exhibit notonly a great stability in vitro but also a surprisingly great stabilityin vivo so that a release or an exchange of the ions, not covalentlybound in the complexes—toxic in themselves—within the time, in which thenew contrast media again are completely excreted now takes placeextremely slowly.

In general, the agents according to the invention for use as NMRdiagnostic media was dosed in amounts of 0.0001-5 mmol/kg, preferably0.005-05 mmol/kg. Details of the use are discussed, for example, in H.J. Weinmann et al., Am. J. of Roentgenology 142, 619 (1984).

Further, the complex compounds according to the invention canadvantageously be used as susceptibility reagents and shift reagents forthe in vivo NMR spectroscopy.

Because of their favorable radioactive properties and of the goodstability of the complex compounds contained in them, the agentsaccording to the invention are also suitable as radiodiagnostic media.Details of their use and dosage are described, e.g., in “Radiotracersfor Medical Applications,” CRC Press, Boca Raton, Fla.

Another imaging method with radioisotopes is positron emissiontomography, which uses positron-emitting isotopes such as, for example,⁴³Sc, ⁴⁴Sc, ⁵²Fe, ⁵⁵Co and ⁶⁸Ga (Heiss, W. D.; Phelps, M. E.; PositronEmission Tomography of Brain, Springer Verlag Berlin, Heidelberg, NewYork 1983).

The compounds according to the invention can also be used in radioimmunoor radiation therapy. It differs from the corresponding diagnosis onlyby the amount and type of isotope used. In this case, the claim is todestroy tumor cells by energy-rich shortwave radiation with a smallestpossible range. Suitable beta-emitting ions are, for example, ⁴⁶Sc,⁴⁷Sc, ⁴⁸Sc, ⁷²Ga, ⁷³Ga and ⁹⁰Y. Suitable alpha-emitting ions exhibitinga short half-life are, for example, ²¹¹Bi, ²¹²Bi, ²¹³Bi and ²¹⁴Bi, and²¹²Bi is preferred. A suitable photon- and electron-emitting ion is¹⁵⁸Gd, which can be obtained from ¹⁵⁷Gd by neutron capture.

If the agent according to the invention is intended for the use in thevariant of radiation therapy proposed by R. L. Mills et al. [Nature Vol.336 (1988), p. 787], the central ion has to be derived from a Moessbauerisotope such as, for example, ⁵⁷Fe or ¹⁵¹Eu.

In the in vivo application of the therapeutic agents according to theinvention, they can be administered together with a suitable carriersuch as, for example, serum or physiological common salt solution andtogether with another protein such as, for example, human serum albumin.In this case, the dosage depends on type of cellular impairment, themetal ion used and the type of method, e.g., brachytherapy.

The therapeutic agents according to the invention are administeredparenterally.

Details of the use of radiotherapeutic agents are discussed, e.g., in R.W. Kozak et al. TIBTEC, October 1986, 262.

The agents according to the invention, because of their outstandingwater solubility and because of the low osmotic pressure of theirconcentrated aqueous solutions, are excellent x-ray contrast media, andit is to be especially emphasized that with them inbiochemical-pharmacological studies no indications can be perceived ofthe known anaphylactic-type reactions from the iodine-containingcontrast media. Because of the favorable absorption properties in theranges of higher tube voltages they are especially valuable for digitalsubtraction techniques.

In general, the agents according to the invention for use as x-raycontrast media are dosed analogously to, e.g., meglumine diatrizoate inamounts of 0.1-5 mmol/kg, preferably 0.25-1 mmol/kg.

Details of the use of x-ray contrast media are discussed, for example,in Barke, Roentgenkontrastmittel [X-ray Contrast Media], G. Thieme,Leipzig (1970) and P. Thurn, E. Buecheler—Einfuehrung in dieRoentgendiagnostik [Introduction to X-ray Diagnosis], G. Thieme,Stuttgart, New York (1977).

Altogether it has been possible to synthesize new complexing agents,metal complexes and metal complex salts, which open up new possibilitiesin diagnostic and therapeutic medicine. Especially the development ofnovel imaging processes in medical diagnosis makes this developmentappear desirable.

Without further elaboration, it is believed that one skilled in the artcan, using the preceding description, utilize the present invention toits fullest extent. The following preferred specific embodiments are,therefore, to be construed as merely illustrative, and not limitative ofthe remainder of the disclosure in any way whatsoever.

In the foregoing and in the following examples, all temperatures are setforth uncorrected in degrees Celsius and unless otherwise indicated, allparts and percentages are by weight.

The entire disclosure of all applications, patents and publications,cited above and below, and of corresponding German application P 40 09119.8 are hereby incorporated by reference.

The purpose of following examples is to provide a more detailedexplanation of the object of the invention.

EXAMPLE 1 a)10-(2,3,4-Trihydroxybutyl)-1,4,7-triscarboxymethyl-1,4,7,10-tetraazacyclododecane

10.0 g (28.87 mmol) of1,4,7-triscarboxymethyl-1,4,7,10-tetraazacyclododecane (DO3A) isdissolved in 40 ml water and the pH is adjusted to 13 with 5 normalsodium hydroxide solution. A solution of 6.24 g (43.30 mmol) of2-(2,2-dimethyl-1,3-dioxolan-4-yl)-ethylene oxide (DE 3 150 917) in 10ml of dioxane is added and stirred for 24 hours at room temperature. Itis diluted with 60 ml of water and extracted three times with 50 ml ofether. The aqueous phase is brought to pH 2 with 10% hydrochloric acidand concentrated by evaporation. The residue is dissolved in some waterand poured on a cation exchange column (IR 120). After rinsing withwater, the ligand is eluted with 0.5 normal aqueous ammonia solution.The fractions are concentrated by evaporation, the ammonium salt istaken up with a little water and poured over an anion exchange column(IRA 67). It is first washed with water and then eluted with 0.5 normalaqueous formic acid.

It is concentrated by evaporation in a vacuum, the residue is dissolvedin a little hot methanol and acetone is added, and the title compound iscrystallized out.

11.31 g (87% of theory) of white powder is obtained, which dissolves inthe air (according to analysis 11.1% water).

Analysis: (corrected for water) Cld: C 47.99 H 7.61 N 12.44 O 31.97 Fnd:C 47.93 H 7.67 N 12.40

b) Gadolinium Complex of10-(2,3,4-trihydroxybutyl)-1,4,7-triscarboxymethyl-1,4,7,10-tetraazacyclododecane

10.0 g (22.2 mmol) of the compound obtained according to 1a) isdissolved in 60 ml of deionized water and 4.02 g (11.1 mmol) ofgadolinium oxide is added. It is heated to 90° C. for 3 hours.

The cooled solution is stirred with 2 ml each of acid ion exchanger (IR120) and 2 ml of basic exchanger (IRA 410) for 1 hour at roomtemperature. It is filtered from the exchanger and the filtrate isbriefly boiled with activated carbon.

After filtering and freeze-drying, 12.76 g (95% of theory) of a whiteamorphous powder is obtained (12.3% water according to analysis).

Analysis: (corrected for water) Cld: C 35.73 H 5.17 N 9.26 O 23.8 Gd25.99 Fnd: C 35.68 H 5.24 N 9.21 Gd 25.93

EXAMPLE 2 a)10-(6-Hydroxy-2,2-dimethyl-1,3-dioxepan-5-yl)-1,4,7-tris(p-toluenesulfonyl)-1,4,7,10-tetraazacyclododecane

50 g (78.76 mmol) of4,7,10-tris(p-toluenesulfonyl)-1,4,7,10-tetraazacyclododecane and 13.63g (94.51 mmol) of 4,4-dimethyl-3,5,8-trioxabicyclo-[5.1.0]-octane aredissolved in 300 ml of dimethylformamide and heated in an autoclave for24 hours to 170° C. It is evaporated to dryness and the residue ischromatographed on silica gel (mobile solvent: methylenechloride/hexane/acetone: 10/5/1). The main fractions are concentrated byevaporation and recrystallized from methyl tert-butyl ether/methanol.Yield: 52.76 g (86% of theory) of a cream-colored powder.

Analysis: Cld: C 55.51 H 6.47 N 7.19 S 12.35 Fnd: C 55.46 H 6.52 N 7.18S 12.32

b)10-(6-Hydroxy-2,2-dimethyl-1,3-dioxepan-5-yl)-1,4,7-tetraazacyclododecane

50 g (64.19 mmol) of the title compound from example 2a) is suspended in800 ml of liquid ammonia/400 ml of tetrahydrofuran and cooled to −35° C.8.9 g (1.28 mol) of lithium is added within 30 minutes and stirred at−35° C. for 8 hours. The excess lithium is destroyed by careful additionof methanol. The ammonia gas is carefully allowed to evaporate and thenevaporated to dryness. The residue is taken up with 200 ml of 4 normalsodium hydroxide solution and extracted three times with 400 ml of hottoluene. The organic phases are dried on potassium hydroxide pellets andthen concentrated by evaporation in a vacuum. The remaining oil ischromatographed (mobile solvent: methanol/water/conc. ammoniasolution=10/1/1). 8.53 g (42% of theory) of a light yellow oil isobtained, which solidifies when left standing.

(8.1% water according to analysis).

Analysis: (corrected for water): Cld: C 56.93 H 10.19 N 17.71 Fnd: C56.88 H 10.15 N 17.64

c)10-(1-Hydroxymethyl-2,3-dihydroxypropyl)-1,4,7-triscarboxymethyl-1,4,7,10-tetraazacyclododecane

8 g (25.28 mmol) of the title compound from example 2b) is dissolved in50 ml of water and 14.05 g (101.12 mmol) of bromoacetic acid is added.The pH is brought to 9.5 with 6normal potassium hydroxide solution.

It is heated to 50° C. and the pH is kept between 9.5-10 by addition of6 n potassium hydroxide solution. After 12 hours of stirring at 50° C.it is cooled in an ice bath, adjusted to pH 2 with concentratedhydrochloric acid and evaporated to dryness in a vacuum.

The residue is dissolved in a little water and poured onto a cationexchange column (IR 120). After rinsing with water, the ligand is elutedwith 0.5 normal aqueous ammonia solution. The fractions are concentratedby evaporation, the ammonium salt is taken up with a little water andpoured onto an anion exchange column (IRA 67). It is washed first withwater and then eluted with 0.5 normal aqueous formic acid. It isconcentrated by evaporation in a vacuum, the residue is dissolved in alittle hot methanol and acetone is added. After cooling in an ice bath,the title compound is crystallized out.

Yield: 8.56 g (69% of theory) of a hygroscopic solid, 9.1% of wateraccording to analysis).

Analysis: (corrected for water): Cld: C 51.42 H 7.81 N 11.42 Fnd: C51.37 H 7.86 N 11.37

d) Gadolinium Complex of10-(1-hydroxymethyl-2,3-dihydroxypropyl)-1,4,7-triscarboxymethyl-1,4,7,10-tetraazacyclododecane

8 g (16.31 mmol) of the title compound from example 2c) is dissolved in50 ml of deionized water and 2.96 g (8.15 mmol) of gadolinium oxide isadded. It is heated for 3 hours to 90° C. The cooled solution is stirredwith 2 ml each of acidic ion exchanger (IR 120) and 2 ml of basicexchanger (IRA 410) for 1 hour at room temperature. The exchanger isfiltered off and the filtrate is briefly boiled with activated carbon.After filtration and freeze-drying, 9.99 g (95% of theory) of anamorphous powder (7.8% of water according to analysis) is obtained.

Analysis: (corrected for water): Cld: C 39.12 H 5.47 N 8.69 Gd 24.39Fnd: C 39.07 H 5.51 N 8.61 Gd 24.32

EXAMPLE 3 Dysprosium Complex of10-(2,3-4-trihydroxybutyl)1,4,7-triscarboxymethyl-1,4,7,10-tetraazacyclododecane

Analogously to the instructions of 1b) the desired dysprosium complex isobtained by starting from 1a) with dysprosium oxide instead ofgadolinium oxide.

Analysis: (corrected for water): Cld: C 35.44 H 5.12 N 9.19 Dy 26.64Fnd: C 35.38 H 5.19 N 9.13 Dy 26.59

EXAMPLE 4 Bismuth Complex of10-(2,3-4-trihydroxybutyl)1,4,7-triscarboxymethyl-1,4,7,10-tetraazacyclododecane

Analogously to the instructions of 1b) the corresponding bismuth complexis obtained by starting from 1a) with bismuth oxide instead ofgadolinium oxide.

Analysis: (corrected for water): Cld: C 32.93 H 4.76 N 8.54 Bi 31.84Fnd: C 32.87 H 4.81 N 8.49 Bi 31.78

EXAMPLE 5 Ytterbium Complex of10-(1-hydroxymethyl-2,3-dihydroxypropyl)-1,4,7-triscarboxymethyl-1,4,7,10-tetraazacyclododecane

Analogously to the instructions of 2d) the corresponding ytterbiumcomplex is obtained by starting from 2c) with ytterbium oxide instead ofgadolinium oxide.

Analysis: (corrected for water): Cld: C 34.84 H 5.04 N 9.03 Yb 27.89Fnd: C 34.79 H 5.10 N 9.01 Yb 27.83

EXAMPLE 6 Lutetium Complex of10-(1-hydroxymethyl-2,3-dihydroxypropyl)-1,4,7-triscarboxymethyl-1,4,7,10-tetraazacyclododecane

Analogously to the instructions of 2 d) the corresponding lutetiumcomplex is obtained by starting from 2 c) with lutetium oxide instead ofgadolinium oxide.

Analysis: (corrected for water:) Cld: C 34.73 H 5.02 N 9.00 Lu 28.11Fnd: C 34.67 H 4.96 N 8.96 Lu 28.06

EXAMPLE 7 Europium¹⁵¹ Complex of10-(1-hydroxymethyl-2,3-dihydroxypropyl)-1,4,7-triscarboxymethyl-1,4,7,10-tetraazacyclododecane

Analogously to the instructions of 2d) the corresponding europium¹⁵¹complex is obtained by starting from 2c) with europium oxide (¹⁵¹Eu₂O₃)instead of gadolinium oxide.

Analysis: (corrected for water:) Cld: C 36.06 H 5.21 N 9.35 Eu 25.35Fnd: C 36.01 H 5.29 N 9.30 Eu 25.29

EXAMPLE 8 Manganese(II) Complex of10-(1-hydroxymethyl-2,3-dihydroxypropyl)-1,4,7-triscarboxymethyl-1,4,7,10-tetraazacyclododecaneas sodium salt

10 g (22.2 mmol) of the title compound from example 2c) is dissolved in80 ml of deionized water and 2.55 g (22.2 mmol) of manganese(II)carbonate is added. It is heated for 3 hours to 90° C.

The cooled solution is stirred for 1 hour with 10 ml of weakly acidicion exchanger (AMB 252c) at room temperature. It is filtered off fromthe exchanger.

The filtrate is adjusted to pH 7.2 with 2N sodium hydroxide solution andfreeze-dried.

Yield: 10.75 g (93% of theory) of a colorless amorphous powder.

(6.3% of water according to analysis).

Analysis: (corrected for water:) Cld: C 41.15 H 5.95 N 10.66 Mn 10.46 Na4.38 Fnd: C 41.08 H 6.03 N 10.58 Mn 10.41 Na 4.43

EXAMPLE 9 Production of a Solution of Indium-111 Complex of10-(1-hydroxymethyl-2,3-dihydroxypropyl)-1,4,7-triscarboxymethyl-1,4,7,10-tetraazacyclododecane

A solution of 100 micrograms of the complexing agent, described inexample 2c), in 5 ml of a mixture of 150 mmolar common salt solution and150 mmolar sodium acetate solution (pH 5.8) is mixed with 4.5 ml ofindium-111 chloride solution (0.01 mmolar) in 1 ml of 0.15 nhydrochloric acid and heated for 1 hour to 80° C. Then the solution isbrought to a pH of 7.2 by addition of 0.1 n sodium hydroxide solution.The solution was sterilized by filtration and freeze-dried. The residueis taken up in physiological common salt solution and then represents apreparation suitable for radiodiagnosis.

EXAMPLE 10 Production of a Solution of Gadolinium Complex of1-(1-hydroxymethyl-2,3-dihydroxypropyl)-4,7,10-triscarboxymethyl-1,4,7,10-tetraazacyclododecane

322.39 g (=0.5 mol) of the compound described in example 2d) isdissolved in 600 ml of water pro injectione (p.i.). After addition of1.5 g of monohydrate of the calcium trisodium salt of DPTA, CaNa₃DPTAand 1.21 g of trishydroxymethylaminomethane a pH of 7.0 is adjusted withdilute hydrochloric acid and water p.i. is added to produce 1000 ml. Thesolution is ultrafiltered, poured into bottles and heat-sterilized.

EXAMPLE 11 Production of a Solution of the Yttrium-90 complex of1-(1-hydroxymethyl-2,3-dihydroxypropyl)-4,7,10-triscarboxymethyl-1,4,7,10-tetraazacyclododecane

5 microliters of a Y-90 solution (2 microcuries) is added to a solutionof 10 micromol of the compound described in example 2d) in 90microliters ammonium acetate buffer (pH 6.0) and the mixture isincubated for 30 minutes at 37° C. 10 micromol of calcium trisodium saltof DTPA is added and, after ultrafiltration a preparation suitable forradiotherapy is obtained.

EXAMPLE 12 Production of a Solution of the Gadolinium Complex of10-2,3,4-trihydroxybutyl)-1,4,7-triscarboxymethyl-1,4,7,10-tetraazacyclododecane

967.5 g of the complex described in example 1b) is suspended in 500 mlof bidistilled water. It is brought to pH 7.3 by addition of 1.89 g ofsodium bicarbonate, 162.3 mg of CaNa₂EDTA is added and with heatingbidistilled water is added to make a volume of 1 liter. After filtrationthrough a pore size of 0.22 microns, the solution is poured intoMultivials and sterilized for 20 minutes at 120° C. A contrast mediumfor x-ray diagnosis is obtained.

EXAMPLE 13 Example for an NMR-Diagnostic In Vivo Examination

Demonstration of a Cerebral Infarction in the Rat by the GadoliniumComplex of10-(1-hydroxymethyl-2,3-dihydroxypropyl)-1,4,7-tris-carboxymethyl-1,4,7,10-tetraazacyclododecane(example 2d) with a dose of 0.1 and 0.3 mmol of Gd/ka

The experimental animal was a female Wistar rat weighing 200 g. For theinduction of the cerebral infarction, the animal was first anesthetized.After intravenous injection of Bengal red with a dose of 20 mg/kg, anarea of the brain about 0.7 cm in diameter near the bregma point wasirradiated through the cranium with light of wavelength 548 nm (thiscorresponds to the maximum absorption of Bengal red), by which, becauseof the formation of singlet oxygen by a chain of various reactions,there results the aggregation of platelets and thus an infarction in theirradiated region.

The imaging took place in an MRI experimental device of the GeneralElectric company (field strength, 2 teslas). It was performed with aspin echo sequence (TR=400 msec, Ts=20 msec). The layer thickness was 3mm, and 4 averagings were performed respectively.

An axial sectional plane image without contrast medium where the craniumpoints downward was taken. The infraction area is indicated only quiteweakly by a lower signal intensity in comparison to the healthy tissue.In an image of the same rat in the same sectional plane, a clearenhancement (clearly increased signal intensity) in the area ofinfraction and the disturbance of the blood-brain barrier connected withit can be seen in an image taken 1 minute after administration of thecontrast medium (0.1 mmol of Gd/kg). The same animal was administeredanother dose after about 20 minutes, this time in the amount of 0.3 mmolof Gd/kg. In an image along the same sectional plane, an enhancementsubstantially stronger than after the smaller dose is shown clearly 1minute after the administration; moreover the infraction area can bebetter defined.

The preceding examples can be repeated with similar success bysubstituting the generically or specifically described reactants and/oroperating conditions of this invention for those used in the precedingexamples.

From the foregoing description, one skilled in the art can easilyascertain the essential characteristics of this invention, and withoutdeparting from the spirit and scope thereof, can make various changesand modifications of the invention to adapt it to various usages andconditions.

1. A method of performing in vivo NMR spectroscopy in a patientcomprising administering a compound as a susceptibility agent which hasthe formula I_(A)

in which each R¹ is hydrogen or a metal ion equivalent, independent ofone another and R² is butyltriol or a salt thereof with an organic orinorganic base or an amino acid, and at least two of the substitutens R¹are metal ion equivalents.
 2. A method of radiation therapy of a patientcomprising administering as a photon and electron emitting agent, acompound which has the formula I_(A)

in which each R¹ is hydrogen or a metal ion equivalent, independent ofone another and R² is butyltriol or a salt thereof with an organic orinorganic base or an amino acid, and at least two of the substitutens R¹are metal ion equivalents.